Quaternary ammonium steroids

ABSTRACT

QUATERNARY AMMONIUM STEROIDS HAVING ANTI-BACTERIAL ACTIVITY WHICH ARE PREPARED BY THE REACTION OF A DIAZABICYCLOALKENE WITH A HALOGEN SUBSTITUTED STEROID OR A STEROID SUBSTITUTED WITH A DISPLACEABLE RADICAL SUCH AS METHANESULFONYLOXY OR TOLUENESULFONYLOXY.

United States Patent 3,553,211 QUATERNARY AMMONIUM STEROIDS David J.Ellis, Mountain View, and David H. Rammler, Palo Alto, Calif., assignorsto Syntex Corporation, Panama, Panama, a corporation of Panama NoDrawing. Filed Aug. 8, 1968, Ser. No. 751,072 Int. Cl. C07c 173/10 US.Cl. 260-2395 26 Claims ABSTRACT OF THE DISCLOSURE Quaternary ammoniumsteroids having anti-bacterial activity which are prepared by thereaction of a diazabicycloalkene with a halogen substituted steroid or asteroid substituted with a displaceable radical such asmethanesulfonyloxy or toluenesulfonyloxy.

This invention relates to novel quaternary ammonium steroids. Moreparticularly, the present invention relates to quaternary ammoniumsteroids characterized by the following Formula A:

in which R represents a steroid molecule; m is a positive integer of twoto seven; n is a positive integer of two to four; and x is apharmaceutically acceptable anion. The wavy line (i) indicates that theconfiguration between the carbon atom of the steroid molecule and thediazabicycloalkene can be alpha, beta or a mixture of alpha and beta,The broken line indicates resonance in the diazabicycloalkene group.

The quaternary ammonium steroids of the present invention are preparedby a displacement reaction in which a halogen atom, preferably bromo,chloro or iodo, or other radical, such as methanesulfonyloxy orp-toluenesulfonyloxy, on the steroid molecule is displaced by adiazabicycloalkene of Formula B resulting in the quaternary ammoniumsteroids (A).

N (012kb In the preparation of the compounds of the present invention,the reaction between the steroid molecule and the diazabicycloalkenemolecule can be carried out in an organic solvent inert to the reaction,such as benzene, toluene, diglyme, petroleum ether, hexane, pentane,xylene, tetrahydrofuran, and the like, at a temperature of from roomtemperature or lower to the reflux temperature of the reaction mixture.Generally, it is preferable to employ elevated temperatures, such asreflux temperature. To minimize side reactions, the reaction ispreferably run under anhydrous conditions and under an inert atmosphere,such as nitrogen. The proportion of reagents is not critical. Equimolaramounts of the steroid and diazabicycloalkene are suitable andpreferably a slight molar excess of the diazabicycloalkene reagent. Theprogress of the reaction can be followed by thin layer chromatographyand, depending upon the reaction temperature, generally takes from aboutminutes to two hours or more 3,553,211 Patented Jan. 5, 1971 to go tocompletion. In most cases, depending upon the stereochemistry of thesteroid molecule, complete or partial inversion takes place. Generally,when the halo or other displaceable radical is attached to a fullysaturated carbocyclic ring of the steroid molecule, inversion takesplace yielding only one isomer. When the halo or other displaceableradical is attached to an unsaturated carbocyclic ring of the steroid,the reaction may yield a mixture of isomers. The isomers can beseparated using fractional crystallization or chromatography.

Diazabicycloalkenes of Formula B are commercially available or, ifdesired, can be synthesized using the procedure of Oediger et al., Chem.Ber., 99, 2012 (1966) or Oediger and Moller, Angew. Chem. Internat.Edit, 6, No. 1, 76 (1967). The diazabicycloalkenes of Formula B include1,5 diazabicyclo[4.3.0]non-5-ene, 1,5-diazabicyclo [5.4.0]undec 5 ene,1,4diazabicyclo[3.3.0]oct-4-ene, 1,6-diazabicyclo[5.3.0]dec-6-ene,1,5-diazabicyclo[6.4.0] dodec-S-ene and 1,6-diazabicyclo [7.5.0]tetradec-G-ene.

The term steroid, as used herein, refers to steroids obtained fromnatural sources, synthetically modified natural steroids and steroidsobtained by total synthesis. Preferably, the steroid molecule orbackbone contains at least 17 carbon atoms. The term steroidal halide,as used herein, refers to a halogen substituted steroid. Preferably, abromo, chloro, or iodo substituted steroid containing at least 17 carbonatoms wherein the halogen atom is at one of positions 03, C-6, C 7,C-16, Cl7, C20, C-21, C-22, C-23 or C24. Suitable steroidal startingmaterials include steroidal halides or steroids containing anequivalently functionalized displaceable radical, such asmethanesulfonyloxy or toluenesulfonyloxy of the gonane, estraine,androstane, pregnane, 19-norpregnane, cholestane, sapogenin,stigmastene, alkaloid or ergostane series. Homoand nor-steroids, such asthe A-homo, B- homo, D-homo, A-nor and B-nor steroids are also suitable.

In addition to the radical which is displaced by the diazabicycloalkeneradical, the steroid molecule can have present other radicals orsubstituents. Substiutents or groups which can be present on thesteroidal starting material include hydroxyl, ester groups, ethergroups, oxo, fiuoro, amino, alkyl, thio, silyl and other conventionalsubstituents of the steroid art.

Substitutents on the steroidal starting material which are sensitive tobase, such as ester groups, may be hydrolyzed in part or completelyduring the reaction. The steroidal starting material can be saturated orcontain monoor polyethylenic unsaturation at, for example, C-1, C3, C4,C5, C-6, C7, C-22, C1,4, C4,6, C-5,7, C3,5, C7,22, C5,22, C5,7,22,C1,3,5(10), C1,4,6, C-1,3,5(10), 6,8 and the like.

The steroidal starting materials which are used for preparing the novelquaternary ammonium steroids of the present invention are commerciallyavailable or can be prepared using known methods. Steroidal halidestarting materials are prepared by treating the steroid molecule with ahalogenating agent such as N-bromosuccinimide, N-chlorosuccinimide,triphenylphosphine dibromide, cupric bromide, thionyl chloride,N-bromoacetamide, phosphorus tribromide, phosphorus pentachloride,phosphorus pentabromide, and the like. Methods of halogenation using theforegoing reagents and others are described in the art.

See, for example, US. Patents 2,255,073; 2,262,244; 2,311,050;2,468,859; 2,531,688; 2,568,025; 2,577,226; and 2,673,206; Wagner andZook, Synthetic Organic Chemistry, John Wiley & Sons, Inc., 88 (1953Djerassi, Steroid Reactions, Holden-Day, Inc., 179 (1963); Fieser andFieser, Reagents for Organic Synthesis, John Wiley & Sons, Inc., 161,862, 865-7, 875, 1247 (1967); and Glazier, J. Org. Chem., 27, 4937(1962).

3 Included among the quaternary ammonium steroids of the presentinvention of Formula A above are those of the following Formulas I-XVI:

in which in, n and x are as defined above;

R is 0x0 or the group in which R is hydrogen, hydroxy andpharmaceutically acceptable esters and ethers thereof, acetyl or loweralkyl of one to ten carbon atoms;

R is hydrogen or hydroxy and pharmaceutically acceptable esters andethers thereof; and

Z is a carbon-carbon single bond or a carbon-carbon 45 double bondbetween 0-1 and C2.

IAO/

and

wherein R and R are as defined above.

(VII) wherein R and R are as defined above;

R is a lower alkylene of one to eight carbon atoms; and Z is acarbon-carbon single bond or a carbon-carbon double bond between C5 andC6.

Y 3 NW an -w R Hy d R (Vlll) wherein R and R are as defined above;

R is methyl or ethyl; and

Z is a carbon-carbon single bond or a carbon-carbon double bond betweenC22 and 0-23.

Pd-R R R, R and R are as defined above;

R is hydrogen, methyl, ethyl or propyl;

Z is a carbon-carbon single bond or a carbon-carbon double bond betweenC8 and Cll; and

Z is a carbon-carbon single bond or a carbon-carbon double bond betweenC6 and C7, provided that when Z is a double bond that Z is a doublebond.

wherein R, R R and Z are as defined above.

The expression lower alkyl, as used herein, refers to lower saturatedaliphatic hydrocarbons, branched and straight chain, such as methyl,ethyl, propyl, isopropyl, n-hexyl, n-pentyl, s-butyl, octyl, and thelike. Preferred are the lower alkyl groups of one to ten carbon atomsassociated with naturally occurring sterols, such as cholesterol,campesterol, ergostanol, sitosterol, stigmasterol, clionasterol, and thelike. The expression lower alkylene, as used herein, refers to lowersaturated divalent aliphatic hydrocarbons, branched and straight chain,such as methylene, ethylidene, propylidene, isopropylidene, butylidene,pentamethylene, trimethylene, tetramethylene, and dimethylene of one toeight carbon atoms.

The expression pharmaceutically acceptable anion, as used herein, refersto a pharmaceutically acceptable anion of an inorganic or organic acidconventionally used in the pharmaceutical art derived from acids, suchas sulfuric, hydrochloric, hydrobromic, hydroiodic, nitric, phosphoric,lactic, benzoic, methanesulfonic, p-toluenesulfonic, salicyclic, acetic,propionic, maleic, tartaric, citric, cyclohexylsulfamic, succinic,nicotinic, ascorbic acids, and the like.

The expression pharmaceutically acceptable esters and ethers thereof, asused herein, refers to pharmaceutically acceptable esters and ethersconventionally employed in the steroid pharmaceutical art. Typical ofthese esters and ethers are acetate, propionate, butyrate,trimethylacetate, valerate, methylethylacetate, caproate,t-butylacetate, 3-methylpentanoate, enanthate, caprylate,triethylacetate, pelargonate, decanoate, undecanoate, benzoate,phenylacetate, diphenylacetate, cyclopentylpropionate, methoxyacetate,aminoacetate, diethylaminoacetate, trichloroacetate, ,8chloropropionate, bicyclo[2.2.2]octane-l'-carboxylate, adamantoate,dihydrogen phosphate, dibenzyl phosphate, sodium ethyl phosphate, sodiumsulfate, sulfate, tetrahydropyran-2-yl ether, tetrahydrofuran-2-ylether, 4-methoxytetrahydropyran-4'-yl ether, methyl ether, ethyl ether,cyclopentyl ether, cyclohexyl ether, propyl ether, and the like.

The quaternary ammonium steroids of the present invention are effectiveagainst bacteria and fungi, such as Bacillus subtilis, Staphylococcusaureus, Escherichia coli, Proteus vulgaris, Salmonella paratyphi,Pseudomonas aeruginas, Shigclla sonnei, Microsporum gypseum, Candido,albicans, T richophytom mentagrophytes, Streptococcus pyogenes,Diplococcus pneumonia, N. aster-aides, Cr. neoformans and S. schenshii.The novel compounds are effective in controlling bacteria and fungiWithin a wide dosage range. Depending upon such factors as the degreeand severity of infection, the type of bacteria or fungi, the compoundsof the present invention are biocidal or biostatic. The compounds of thepresent invention can be used in solutions, sprays, and the like, fordisinfecting inanimate objects or can be used topically in the form ofointments, creams, solutions, sprays, and the like, for treatingbacterial infections of animals.

The following examples are provided to illustrate the present invention.

EXAMPLE 1 (A) To a mixture of 1 g. of 3/3-hydroxyandrost-5-en- 17-one inml. of benzene is added 1.5 g. of phosphorus tribromide. The reactionmixture is refluxed for about one hour under anhydrous conditions andthen cooled and poured into water. The organic layer is washed withWater several times, dried over sodium sulfate and evaporated to drynessto yield 3 3-bromoandrost-5-en-17-one.

By repeating the above procedure using phosphorous trichloride inchloroform, there is obtained 3fi-chloroandrost-5-en-17-one.

Alternatively, thionyl bromide and thionyl chloride in a dry ethersolvent at about room temperature can be used to prepare 3-bromo and3-chloro derivatives from the corresponding 3-hydroxyl.

(B) A mixture of 2 g. of 3/8-bromoandrost-5-en-17-one, ml. of xylene and10 molar equivalents of 1,5-diazabicyclo[4.3.0]non-5-ene is refluxedunder nitrogen for about two hours with stirring. The reaction mixtureis cooled, concentrated under vacuum and the residue washed with hexane.The insoluble portion of the residue is dis solved in dilute aqueoussodium chloride and extracted with chloroform. The chloroform extractsare combined and evaporated to dryness to yield3,B-[5'-(1',5'-diazabicyclo[4.3.0]nonenyl)]androst 5 en-17-one bromide(XIII; R is oxo, m is three, 11 is three, x is bromo).

By using 3fl-chloroandrost-5-en-17-one as the starting material, thereis obtained 3,8-[5'-(1,5-diazabicyclo- [4.3.0] nonenyl)androst-5-en-17-one chloride.

EXAMPLE 2 The procedure of Example 1A is repeated using 3,8-hydroxy-5u-androstan-17-one and 3B-hydroxyandrost-4- ene as the startingmaterial and there is obtained 3abromo-5a-androstan-l7-one and3fl-bromoandrost-4-ene. The thus-obtained steroidal bromides are treatedaccording to the procedure of Example IE to afford 3,8-[5'-(1,5-diazabicyclo[4.3.0]nonenyl)] 5a androstan-17- one bromide and amixture of 3}3-[5'-(1',5'-diazabicyclo- [4.3.0]nonenyl)]androst-4-eneand 3a-[5-(1,5'-diazabicyclo [4.3.0] nonenyl)] androst-4-ene.

EXAMPLE 3 (A) A mixture of 2 g. of 35 acetoxyandrost-5-en-17- one, 1molar equivalent of N-bromosuccinimide, and 100 m1. of hexane is stirredunder nitrogen and irradiated with a 375 watt bulb for about 15 minutes.The reaction mixture is poured into ice-water, filtered and the solidwashed with hexane. The filtrate is dried and evaporated to dryness toyield 7fi-bromo-3fl-acetoxyandrost-S-en-17-one.

(B) The product of part A is dissolved in hexane and 5 molar equivalentsof 1,5-diazabicyclo[4.3.0]non-5-ene added. The reaction mixture isrefluxed under nitrogen for 1.5 hours. The reaction mixture is thenevaporated in vacuo, the residue triturated with cold, fresh hexane,decanted and the residue dissolved in methanol. The methanol solution isslowly filtered through a column of ionexchange resin (OH form) withmethanol. The eluate is then passed through a column of ion-exchangeresin (Clform) with methanol and the eluate evaporated in vacuo. Theresidue is dissolved in chloroform, washed with saturated aqueous sodiumchloride, and evaporated. The residue is dissolved in methanol, treatedwith charcoal,

filtered and the filtrate concentrated by boiling. Benzene is added tothe concentrate, heated, cooled and filtered to yield a mixture of7a-[5-(1,5'-diazabicyclo[4.3.0]nonenyl) ]-3,B-hydroxyandrost en-l7-onechloride and 75- [5'-(l',5'-diazabicyclo[4.3.0]nonenyl)] 3Bhydroxyandrost-5-en-17-one chloride which can be separated by fractionalcrystallization.

EXAMPLE 4 A mixture of 2 g. of cholesteryl acetate, 1 g. of N-bromosuccinimide and 80 ml. of hexane is stirred under nitrogen andirradiated with a 375 watt photoflood lamp for minutes. The mixture iscooled in ice and filtered. To the filtrate is added ml. of1,5-diazabicyclo[5.4.0]- undec-S-ene and the resulting mixture isrefluxed under nitrogen for 1.5 hours. The reaction mixture is cooled,concentrated and the residue washed with hexane. The solid obtained isredissolved in methanol, and filtered through a column of ion-exchangeresin (OH- form) and then through ion-exchange resin (Clform). Theeluate is evaporated in vacuo and the residue obtained redissolved inchloroform and Washed with saturated aqueous sodium chloride. Thechloroform solution is evaporated in vacuo, the residue evaporated witha little more chloroform, and the solution filtered. The filtrate isevaporated in vacuo, a little methanol added followed by benzene and thesolution concentrated by boiling to yield a mixture of7a-[5'-(1',5'-diazabicyclo[5.4.0]undecenyl)]-3fl-hydroxycholest-5-enechloride and 7B-[5'- (l',5'-diazabicyclo[5.4.0]undecenyl)] 3/3hydroxycholest-S-ene chloride.

EXAMPLE 5 The procedure of Example 3B is repeated using 75-bromo-3B-acetoxycholest-5-ene at the starting material in place of7,6-bromo-3fl-acetoxyandrost-S-en-17-one, there is obtained a mixture of7ot-[5'-(l',5-diazabicyclo[4.3.0]- nonenyl)]-3;3-hydroxycholest-5-enechloride and 7B-[5'- I (1',5' diazabicyclo[4.3.0]nonenyl)] 3,6hydroxycholest-S-ene-chloride.

By using 7B-bromocampesterylacetate and 7fi-bromositosterylacetate(prepared according to US. Pat. 2,568,025) as the starting material inthe process of Example 38, there is obtained a mixture of 7u-[5'-(l',5'-diazabicyclo[4.3.0]nonenyl)]campest 5 en-SB-ol chloride and713-[5'-(l',5 diazabicyclo [4.3.0]nonenyl)]- campest-5-en-3/3-olchloride and a mixture of 7uc-[5'-(1',5-diazabicyclo[4.3.0]-nonenyl)]sitost 5 en-3B-ol chloride and7B-[5-(1,5'-diazabicyclo[4.3.0]nonenyl)]- sitost-5-en-3fl-ol chloride,respectively.

In the case of Examples 3, 4 and 5, in the work-up of the reactionproduct, the filtering through a chloride form ion-exchange column canbe eliminated and in which case the final products obtained bear abromide anion instead of the chloride anion.

EXAMPLE 6 A mixture of 2 g. of 3fl-chlorocholest-4-ene, 10 molarequivalents of 1,5-diazabicyclo[4.3.0]non-5-ene and 150 ml. of xylene isrefluxed under nitrogen with stirring for two hours. The reactionmixture is evaporated in vacuo and the residue washed with hexane. Theinsoluble portion is mixed into dilute aqueous sodium chloride andextracted with chloroform which is evaporated in vacuo to yield 3/3-[5'(l',5' diazabicyclo[4.3.0]nonenyl)]- cholest-4-ene chloride.

By using cholesteryl iodide as the steroidal halide starting material inthe procedure of this example, there is obtained -[5' (1',5diazabicyclo[4.3.0]nonenyl)]- cholest-S-ene iodide. Cholesteryl idodidecan be obtained as described in US. Pat. 2,311,050. 3,8-Chlorocholest-4-one is prepared by treating 3fl-hydroxycholest-4-ene with thionylchloride according to the procedure of Example 1A. Similarly,3fi-bromoandrost-4-ene is obtained by treating androst-4-en-3fl-ol withthionyl bromide.

8 EXAMPLE 7 The procedure of Example 6 is repeated with the exceptionthat an equivalent amount of the 3-bromo derivative of diosgenin is usedas the steroidal halide starting material to yield 3/3 5' (1',5diazabicyclo[4.3. 0] nonenyl) ]spirost-S-ene bromide. In the samemanner, the 3-bromo derivative of tigogenin is converted into 3,8-[5'-(l,5-diazabicyclo[4.3.0]nonenyl)] 5a spirostane bromide.

7,8-bromodiosgenin acetate is subjected to the procedure of Example 3Band there is obtained a mixture of7a-[5'-(1,5-diazabicyclo[4.3.0]nonenyl)]spirost 5 ene chloride and7,8-[5' (l',5'-diazabicyclo[4.3.0]nonenyl) spirost-S-ene chloride. Byrepeating this procedure with exception of omitting the chloride formion-exchange filtration in the work-up, there is obtained thecorresponding bromide.

The steroidal halide (3-bromo derivative of diosgenin and 3-bromoderivative of tigogenin) can be prepared by treatment of the 3-hydroxysteroid with phosphorus tribromide and thionyl bromide, respectively, asdescribed in Example 1A. The 7-bromo steroidal halide can be obtained bytreating diosgenin acetate with a halogenating agent such asN-bromosuccinimide according to the procedure of US. Pats. 2,531,688 or2,577,226.

EXAMPLE 8 Ergosterol is subjected to the procedure of Example 1A and Bin place of 3fl-hydroxyandrost-5-en-17-one to yield 35-[-5-(1,5'diazabicyclo[4.3.0]nonenyl)]ergosteryl bromide and the correspondingchloride. Similarly, ergosteryl iodine (obtained according to US. Pat.2,3l1,- 050) is converted into 3fl-[5'-(1',5'-diazabicyclo[4.3.0]nonenyl) ]ergosteryl iodide.

Ergosteryl acetate is hydrogenated using a palladiumcharcoal catalystuntil the theoretical amount of hydrogen is consumed to obtain the5,6,22,23-tetrahydro derivative which is subjected to the process ofExample 4 to yield a mixture of6a-[5-(l',5-diazabicyclo[4.3.0]nonenyl)]- 5u-cholest-7-ene chloride and6;8-[5'-(1',5-diazabicyc1o [4.3.0] nonenyl) ]-5a-cholest-7-ene chloride.

EXAMPLE 9 Estrone methyl ether is treated withmethylenetriphenylphosphorane to obtain the l7-methylene derivativethereof which is treated with diborane followed by hydrogen peroxide bythe method of US. Pat. 3,385,849 to yield 3-methoxy-l7 6-hydroxymethylestra-1,3,5(10)- triene which is reacted with phosphorustribromide to yield 1713 bromomethyl-3-methoxyestra-1,3,5(10)-triene.The thus-obtained steroidal halide is subjected to the procedure ofExample IE to yield 20;8-[5'-(1',5'-diazabicyclo[4.3. 0]nonenyl)] 3methoxyestra 1,3,5 10)- triene bromide (X; R is methoxy, R is methyl, Ris methylene, Z and Z are single bond, in is three, n is three).

The 3-methyl ether of 3,20-dihydroxy-19-norpregnal,3,5(l0)-triene istreated with phosphorus tribromide to afford the 20-bromo derivativewhich is subjected to the procedure of Example IE to yield a mixture ofthe on and ,8 isomer of20-[5'-(1',5'-diazabicyclo[4.3.0]nonenyl)]-3-methoxy-19-norpregna-1,3,5(10)-triene.

EXAMPLE 10 The following steroidal chlorides:25-chloro-27-norcholest-5-en-3,B-ol; 25-chloro-27-norcholest-5-en 3 one;and 24-chloro-3fi-acetoxy-27-norcholest-5-ene (prepared according toU.S. 2,673,206) are used as the starting material in the process ofExample IE to yield: 25-[5'- (l,5'-diazabicyclo[4.3.0]nonenyl)] 27norcholest-S- en-3fi-ol chloride; 25-[5 (1',5'diazabicyclo[4.3.0]nonenyl)]-27-norcholest-5-en-3-one chloride; and24-[5'-(l, 5-diazabicyclo[4.3.0] nonenyl) ]-27-norcholest-5 en 3,8- 01chloride.

9 EXAMPLE 11 The procedure of Example 1A and B is repeated using anequivalent amount of a-spinasterol as the starting material to yield35-[5-(l,5-diazabicyclo[4.3.0]nonenyl)] a-spinasteryl bromide. In thesame manner, androst-S- ene-35,17{3-diol 17-benzoate anda-androstan-35,17{3-diol 17-benzoate are converted into 36-[5'(1',5-diazabicyclo [4.3.0]nonenyl) ]-androst-5-en-17/3-ol bromideand 3,8- [5'-(1',5-diazabicyclo] [4.3.0]none-nyl)] 50c androstan-1713-01 bromide.

EXAMPLE 12 The process of Example 4 is repeated using an equivalentamount of androst-5-en-l7B-ol 17-acetate and androst-5ene-3,B-17fl-diol3,17-diacetate as the starting material to yield a mixture of the a andB isomers of 7- [5(l,5'-diazabicyclo[5.4.0]undecenyl)]androst 5 en-176-01 chloride and a mixture of the or and e isomers of 7-[5 (1,5diazabicyclo[5.4.0]undecenyl)]androst- 5-ene-3,8,17B-diol chloride. Byrepeating this procedure using 1,5-diazabicyclo-[4.3.0]non-5-ene inplace of 1,5- diazabicyclo[5.4.0]undec-S-ene, the corresponding 1,5-diazabicyclo[4.3.0]nonenyl chlorides are obtained.

By using an equivalent amount of androst-4-ene-35, 17fl-diol 17-acetatein the procedure of Example 1A and B, there is obtained3-[5'-(1,5-diazabicyclo[4.3.0] nonenyl) androst-4-en-17fl-ol bromide.

EXAMPLE 13 The processes of Example 1A and B are repeated with theexeception of using an equivalent amount of (1) pregn-5-ene3p,20,8-diol3-acetate, (2) 5a-pregnane-3/3, ZOB-diol 3-acetate, (3)pregn-5-ene-3,B,21-diol 3-acetate and (4) 5ot-pregnane-3fi,2l-diol3-acetate as the starting material to yield (1)20-[5'-(1',5-diazabicyclo[4.3.0] nonenyl)]pregn-5-en-3p-ol bromide, (2)20-[5'-(1',5'- diazabicyclo [4.3.0] nonenyl) ]-5a-pregnan-3 8-olbromide, (2) 21-[5' (1,5' diazabicyclo[4.3.0]nonenyl)]pregn- 5-en-3 6-olbromide and (4) 21-[5'-(1',5'-diazabicyclo[4.3.0]nonenyl)]-5a-pregnan-3,8-o1 bromide, respectively.

By using an equivalent amount of (5)17B-hydroxymethylandrost-S-en-313-01 3-acetate,

(6) 17,8hydroxymethyl-5u-androstan-3 5-01 3-acetate,

(7) 20-hydroxymethylpregn-5-en-35-01 3-acetate,

(8) 3 fl-acetoxy-22-hydroxymethyl-23 ,24-bisnorchol-5 ene,

(9) 3[i-acetoxy-22-hydroxymethyl-23,24-bisnor-5acholane,

(10) 3,B-acetoxy-23-hydroxymethyl-24-norchol-5-ene and ( 1 1 3,B-acetoxy-Z3-hydroxymethyl-24-nor-5 a-cholane in place of3fl-hydroxyandrost-5-en-l7-one in the procedures of Example 1A and B,there is obtained (5) -[5-(1',5'-diazabicyclo[4.3.0]nonenyl)1androst-(6) 20-[5'-(1,5'-diazabicyclo[4.3.0]nonenyl)1-Serandrostan-3fl-olbromide,

(7) 22-[5'-(1',5'-diazabicyclo[4.3.0]nonenyl)1-23- norchol-5-en-3b-olbromide,

( 8) 23- [5-( 1,5-diazabicyclo[4.3.0]nonenyl) 1-24- norchol-S-en-Zfi-olbromide,

(9) 23-[5'-(1',5-diazabicyclo[4.3.0]nonenyl)1-24- nor-5u-cholan-3fl-olbromide,

(10) 24-[5'-(1',5-diazabicyclo[4.3.0]nonenyl) ]chol- 5-en-3B-ol bromideand (11) 24-[5'-(1,5-diazabicyclo[4.3.0]nonenyl)1-Sercholan-3B-olbromide, respectively.

The following is a typical procedure which can be used for preparing theabove alcohols.

A solution of 1 g. of 3B-acetoxy-5a-androstane-17B- carboxylic acid in50 ml. of tetrahydrofuran is added over a minute period to a stirredsuspension of 1 g. of lithium aluminum hydride in ml. of anhydroustetrahydrofuran and this mixture is heated at reflux for two hours. Tothis mixture is cautiously added 5 ml. of ethyl acetate and 2 ml. ofwater. Sodium sulfate is next added, the mixture is filtered and thesolid thus collected is washed with hot ethyl acetate. The combinedorganic solutions (filtrate and washings) are then evaporated to yieldl7fi-hydroxymethyl-5u-androstan-3B-o1 3-acetate which may be furtherpurified through recrystallization from acetonezhexane.

In a similar manner,

3fl-acetoxypregn-S-ene-ZO-carboxylic acid,3,8-acetoxy-23,24-bisnorchol-5-ene-22-carboxylic acid,3fl-acetoxy-23,24-bisnor-5a-cholane-22-carboxylic acid,3fi-acetoxy-5a-cholanoic acid,

3B-acetoxychol-5-enoic acid and 3fi-acetoxyandrost-5-en-17/3-carboxylicacid are converted into the corresponding alcohol.

EXAMPLE 14 A mixture of 1 g. of 7,8-[5-(1',5'-diazabocyclo[4.3.0]nonenyl)]cholest-5-en-3,B-ol bromide, 10 ml. of acetic anhydride and 10ml. of pyridine is stirred and left to stand overnight. The reactionmixture is then poured into icewater and the solid which forms iscollected by filtration, washed with water, and dried to yield 78-[5-(l',5'- diazabicyclo [4.3.0] nonenyl) ]cholest-5-en-3;3-oldiacetate.

Similarly, through the use of other carboxylic anhydrides in the aboveprocess, such as propionic anhydride, trifiuoroacetic anhydride, butyricanhydride, benzoic anhydride, trimethylacetic anhydride, and the like,other diacylates can be prepared.

A solution of 1 g. of 7fl-[5-(1,5-diazabocyclo[4.3.0]nonenyl)]cholest-5-en-3,8-ol diacetate in ml. of chloroform is saturatedwith a stream of HCl gas and allowed to stand for about one hour. Thereaction mixture is then evaporated to dryness to yield7B-[5'-(1,5-diazabicyclo[4.3.01nonenyl)]cholest S en-3B-ol 3-acetatechloride which can be purified using chromatography orrecrystallization.

EXAMPLE 15 A mixture of 100 mg. of 24-iodo-5B-cholane, 5 ml. of benzeneand 0.1 g. of 1,5-diazabicyclo[4.3.0]non-5-ene is stirred at about 20for 24 hours. The reaction mixture is then evaporated, the residuedissolved in methanol, and the methanol solution filtered through acolumn of ion-exchange resin (Clcycle) to yield 24[5-(1,5-diazabicyclo4.3 .0] nonenyl) ]-5B-cholane chloride.

EXAMPLE 16 The process of Example 1B is repeated using 163-bromoestr-5(10)-en-17-one, 6tx-chloro 16amethyl-5apregn-Z-ene-17a,21diol-11,20-dione and6wchloro-21-dimethylaminomethyl-A -progresterone as the steroidal halidestarting material to yield 16a-[5-(l,5-diazabicyclo[4.3.0-1nonenyl)]estr-5(10)-en-l7-one bromide, 6B-[5'(1,5'diazabicyclo[4.3.0]nonenyl)]l6u-methyl-5u-pregn-2-ene-l7a,21-diol-l1,20-dione chloride and 6 3-[5-(1',5'-diazabicyclo[4.3.0 ]nonenyl)] 21 dimethylaminomethyl-A-progesterone.

Androst- 1-ene-3 13, 1 7 ,B-diol and 2fi-hydroxymethyl-19-nor-5a-pregnane-20-one are converted into 3fl-chloroandrost-1-en-17fl-oland Zfi-chloromethy l- 19 ROI-5apregnane-ZO-one using thionyl chloridewhich are converted into 3 [5 (1,S diazabicyclo[4.3.0]nonenyl)]androst-1-en-1713-ol chloride and2B-[5'-(1',5'-diazabicyclo[4.3.01nonenyl)]-methyl-19-nor 5a pregnane-20-one chloride.

EXAMPLE 17 The process of Example 4 is repeated using 3B-acetoxypregn-5-en-20'one to yield a mixture of the a and 13 isomers of7-[5'-(1,5-diazabicyclo[5.4.0]undecenyl)] 3fi-hydroxypregn-S-en-ZO-onechloride. By using 1,5-diazabicyclo[4.3.0]non-5-ene in this procedure,there is ob- 1 1 tained 7[5'-(l,5'-diazabicyclo[4.3.0]nonenyl)]-3;3-hydroxypregn-S-en-ZO-onechloride.

EXAMPLE 18 The 3-acetate of N-ergostenol is used as the startingmaterial in the process of Example 3 to yield 6-L5'-(1,5'- diazabicyclo[4.3.0] nonenyl) ]-A -ergostenol chloride. The 3-acetate of N-ergostenolis prepared by treating A ergostenol with acetic anhydride in pyridine.

EXAMPLE 19 The reaction of 60L-bI'O1'1lOChO16SlI3Il6,35-chloroch0lestane, 20-chloropregn-4-en-3-one, 3fl-chloroandrost-l-en-17B-ol 17-acetate, 6fl-Chi0IO-170t-H16thYl B nortestosterone, 3/3-methoxy-7B-chloro B homoestr-5(l0-en- 17-one and3fi-methoxy-6B-chloromethylestr-5 10)-en-17- one with1,5-diazabicyclo[4.3.0]nn-5-ene according to the procedure of Example 6yield 6,8['-(l,5-diazabicyclo [4.3 .0] nonenyl) ]cholestane bromide,3oc- L5 l,5 diazabicyclo[4.3.0]nonenyl)Jcholestane chloride, 20-[5'-(l,5-diazabicyclo[4.3.0]nonenyl)]pregn-4-en 3 one chloride, 35-[5 (1',5'diazabicyclo[4.3.0-]-nonenyl)] androst-l-en-17fi-ol chloride,6-[5-(l,5-diazabicyclo [4.3.0] nonenyl) ]-17a-methyl-B-nortestosteronechloride, 3fi-methoxy 7 [5' (1,5' diazabicyclodiaza[4.3.0]nonenyl)]-B-homoestr-5(10)-en-17-one chloride and 35- methoxy 6B[5'-(1',5'-diazabicyclo[4.3.0]nonenyl)] methylestr-S lO)-en-17-onechloride, respectively.

EXAMPLE 20 (A) A solution of 1 g. of sodium borohydride in 3 ml. ofwater is added to an ice-cooled solution of 1 g. of 3methoxy-19norpregna-1,3,5(l0),l7(20)-tetraen-21-al in 120 ml. ofmethanol and the mixture then allowed to stand for 16 hours at roomtemperature. Excess reagent is decomposed by the addition of acetic acidand the solution is then concentrated to small volume under vacuum anddiluted with water. The product is extracted with ethyl acetate and theextracts are washed with water, dried and evaporated to yield3-methoxy-19-norpregna-1,3,5, (),17(20)-tetraen-21-ol.

(B) The above 2l.-hydroxy-3-methoxy-l9-norpregna- 1,3,5(l0),l7(20)-tetraene is converted into the corresponding 2l-bromocompound which is reacted with 1,5- diazabicyclo[4.3.0]non-5-eneaccording to the procedure of Example 1 to yield21-[5-(1',5'-diazabicyclo[4.3.0] nonenyl)] 3methoxy-19-norpregna-1,3,5(10),17(20)- tetraene bromide.

The 2l-aldehyde used in part A of this example is prepared from thecorresponding l7-keto compound using the procedure of US. Patent3,374,253. Similarly, 3-methoxy-18-methylestra-l,3,5(10),8tetraen-l7-one and3-methoxyestra-1,3,5(l0),6,8-pentaen-l7-one are converted into 3-methoxy 19 nor-18-methylpregna-1,3.5(10),8,17(20)- pentaen-Zl-al and3-methoxy-19-norpregna-l,3,5(l0),6,8, 17(20)-hexaen-21-al which aretreated according to part A and B of this example to yield21[5'-(1',5-diazabicyclo [4.3.0]nonenyl)] 3 methoxy18-rnethyl-19-norpregna- 1,3,5(10),8,17(20)-pentaene bromide and21-[5'-(1,5'- diazabicyclo[4.3.0]nonenyl)] 3 meth0xy-19-norpregna-1,3,5( 10) ,6,8,l7 ()-hexaene bromide.

EXAMPLE 21 The procedure of Example 20 is repeated using 3- methoxy 19norpregna 1,3,5(10)-trien-2l-one, 3- methoxy 18 methyl19-norpregna-l,3,5(10)-trien-2lone and3-methoxy-l9-norpregna-l,3,5(10),8-tetraen-21- one as the startingmaterial to yield 21-[5'-(1,5-diazabicyclo[4.3.0]nonenyl)] 3methoxy-19-norpregna-1,3,5 (10) -triene bromide,2l-[5'-(1,5'-diazabicyclo[4.3.0] nonenyl)] methoxy18-methyl-l9-norpregna-1,3,5(10)- triene bromide and2l-[5-(l,5-dazabicyclo[4.3.0]nonenyl] 3 methoxy .l9-norpregna-l,3,5(lO),8-telraene bromide, respectively.

3 Methoxy l9-norpregna-1,3,5(10)-trien-21-one, 3- methoxy 18methylpregna 1,3,5( l0)-trien-21-one and 3 methoxy l9norpregna-1,3,5(lO),8-tetraen-21-one are obtained by catalytichydrogenation of 3-methoxy-19- norpregna 1,3,5 (l0),17(20)tetraen-Zl-al, 3-rnethoxy- 18 methyl19-norpregna-l,3,5(l0),17(20)-tetraen-2l-al and3-methoxy-l9-norpregna-1,3,5(10),8,17(20)-pentaen- 21-al, respectively,using palladium-on-carbon. See, for example, US. Patent 3,120,518.

EXAMPLE 22 A mixture of 1 g. of 3-methoxyestra-l,3,5(l0)-trien 17,8-01in 5 ml. of pyridine and 0.5 g. of methanesulfonyl chloride is allowedto stand at room temperature for 24 hours and is then diluted with waterand filtered. The solid thus collected is dried and recrystallized fromacetone: hexane to yield 17fi-methanesulfonyloxy-3- methoxyestra-1,3,5(l0)-triene.

By using p-toluenesulfonyl chloride in the above procedure, thecorresponding 17-tosylate is obtained.

The above compounds are reacted with 1,5-diazabicyclo[4.3.0]non-5-eneusing the procedure of Example 18 to yieldl7a-[5-(1,5'-diazabicyclo[4.3.0]nonenyl)]-3-methoxyestra-1,3,5(10)-triene methanesulfonate and 17a- [5'(l,5-diazabicyclo[4.3.0]nonenyl) ]-3-methoxyestra- 1,3,5( 10 -trienep-toluenesulfonate.

EXAMPLE 23 A mixture of 1 g. of 7x-bromo-3,8,l7fldiacetoxy-5aandrostane, ml. of hexane and 4 molarequivalents of 1,5-diazabicyclo[4.3.0]non-5-ene is refluxed undernitrogen for three hours with stirring. The reaction mixture is cooled,concentrated under vacuum and the residue washed with hexane. Theinsoluble portion of the residue is dissolved in aqueous sodium chlorideand extracted with chloroform. The chloroform extracts are combined andevaporated to dryness to yield 7,8-[5-(1,5'-diazabicyclo[4.3.0]nonenyl)]50c androstane 313,175 diol bromide.

The above procedure is repeated using 6-chloro-17,8-(l-cyclohexenyloxy)-B-norandrost-4-en-3-one to yield 6- [5'(l,5-diazabicyclo[4.3.0]nonenyl)]17B-(1'-cyclohexenyloxy)-B-norandrost-4en-3-one chloride.

7a-bromo-3fi,1713-diacetoxy-5a-androstane can be prepared by treating3p,17,8-diacetoxy-Sa-androstan-7-one with lithium tri(t-butoxy)aluminumhydride followed by bromination or chlorination of the thus formedhydroxyl using phosphorus tribromide or trichloride.

EXAMPLE 24 A mixture of 20 g. of 3fl-acetoxyeholest-5-ene, 10 g. ofN-bromosuccinimide and 800 ml. of hexane is rapidly stirred undernitrogen and irradiated with two photofiood lamps (375 watt) for 15minutes. The mixture is stirred in ice, filtered and the solid washedwith hexane. The filtrate and 20 ml. of 1,5-diazabicyclo[4.3.0]non-5-eneis refluxed under nitrogen for 1.5 hours. The reaction mixture is thenevaporated under vacuum and the residue triturated with cold hexanewhich is decanted. The residue is dissolved in methanol and filteredthrough a column of ion-exchange resin (OH form) in methanol. The eluateis then passed through a column of ion-exchange resin (Cl form) inmethanol. The eluate is evaporated under vacuum. The residue isredissolved in chloroform which is washed with water containingsaturated sodium chloride and evaporated under vacuum. The residue isredissolved in methanol, treated with charcoal, filtered throughdiatomaceous earth and the filtrate concentrated to small volume byheating. Benzene is added to the concentrate and the mixture heateduntil cloudy. The mixture is cooled and the crystalline solid filteredoff to yield a mixture of the alpha and beta isomers of7-[5-(l',5'-diazabicyclo- I430 [nonenyl) lcholesl-5-en-3/i-ol chloride.

The quaternary ammonium steroids of the present invention can be namedeither as a derivative of the steroid 13 molecule as above or asderivative of the salt. For example, the quaternary ammonium steroid ofExample 24 which has the following formula:

can be named either7-[5'-(1,5'-diazabicyclo[4.3.0]nonenyl)]-cholest--en-3B-ol chloride or5-( 3'13-hydroxycholest 5' en-7-yl)-1,S-diazabicyclo[4.3.0]n0nenoniumchloride.

What is claimed is:

1. A quaternary ammonium steroid selected from those of the followingFormulas I, II, III and IV:

N) V L (III) (IV) wherein R is the group ('(CHQmw in which m is apositive integer of two to seven; 11 is a positive integer of two tofour; and x is a pharmaceutically acceptable anion;

R is OX0 or the group in which R is hydrogen, hydroxy andpharmaceutically acceptable esters and ether thereof, acetyl or loweralkyl of one to ten carbon atoms;

R is hydrogen or hydroxy and pharmaceutically acceptable esters andethers thereof; and

Z is a carbon-carbon single bond or a carbon-carbon double bond betweenC-1 and C-2.

2. A steroid according to Formula I of claim 1 selected from those ofthe formula:

m a e u s in which R x and Z are as defined therein.

3. A steroid according to Formula II of claim 1 selected from those ofthe formula:

' T .QU...

in which R and x are as defined therein.

5. A compound according to Formula IV of claim 1 selected from those ofthe formula:

in which R R and x are as defined therein.

6. A compound according to claim 2 wherein Z is a single bond.

7. A compound according to claim 2 wherein Z is a single bond and R is0X0.

8. A compound according to claim 3 wherein R is 9. A compound accordingto claim 5 wherein R is 0x0 and R is hydroxy and pharmaceuticallyacceptable esters and ethers thereof.

10. A compound according to claim 5 wherein R is the group and R ishydroxy and pharmaceutically acceptable esters and ethers thereof.

15 11. A compound according to claim in which R is the lower alkyl grouporpwnnros R is hydroxy and x is chloro.

12. A quaternary ammonium steroid selected from those of the Formulas Vand VI:

/N/\'/ KYJ and i O O) R f\% H wherein R "is the group in which m is apositive integer of two to seven; 11 is a positive integer of two tofour; and x is a pharmaceutically acceptable anion; and R is hydrogen ofhydroxy and pharmaceutically acceptable esters and ethers thereof. 13. Asteroid according to Formula V of claim 12 selected from those of theformula:

wherein R is hydroxy and pharmaceutically acceptable esters and ethersthereof.

14. A steroid according to Formula, VI of claim 12 wherein m is three nis three.

15. A quaternary ammonium steroid selected from those of the FormulaVII:

Z (VlI) wherein R is the group 20 2)m I r4. N Xe u a n n a I in which mis a positive integer of two to seven; 12 is a positive integer of twoto four; and x is a pharmaceutically acceptable anion; R is hydrogen orhydroxy and pharmaceutically acceptable esters and ethers thereof;

R is a lower alkylene of one to eight carbon atoms;

and Z is a carbon-carbon single bond or a carbon-carbon double bondbetween C-5 and C-6. 16. A steroid according to claim 15 wherein R ismethylene or ethylene and each of m and n is three.

17. A steroid according to claim 15 selected from those of the formula:

0113 l; N i oI-IMN in which R at and Z are as defined therein.

18. A steroid according to claim 15 selected from those of the formula:

in which R x and Z are as defined therein and p is a positive integer ofone to three.

19. A quaternary ammonium steroid selected from those of the FormulasVIII and IX:

in which m is a positive integer of two or seven; 11 is a positiveinteger of two or four; and x is a pharmaceutically acceptable anion;

R is hydrogen or hydroxy and pharmaceutically acceptable esters andethers thereof;

R is methyl or ethyl; and

Z is a carbon-carbon single bond or a carbon-carbon double bond between022 and C23.

20. A steroid according to Formula VIII of claim 19 selected from thoseof the formula:

in which R is hydroxy and pharmaceutically acceptable esters and ethersthereof and x is as defined therein.

21. A quaternary ammonium steroid selected from those of the Formulas X,XI and XII:

18 wherein t R is the group (CHQmw J Xe N es N in which m is a positiveinteger of two to seven; 21 is a positive integer of two to four; and xis a pharmaceutically acceptable anion; R is hydrogen or hydroxy andpharmaceutically acceptable esters and ethers thereof; R is a loweralkylene of one to eight carbon atoms; R is hydrogen, methyl, ethyl orpropyl; Z is a carbon-carbon single bond or a carbon-carbon double bondbetween C 8 and Cl1; and Z is a carbon-carbon single bond or acarbon-carbon double bond between C-6 and C-7, provided that when Z is adouble bond that Z is a double bond. from a 50:1 dilution to a 50011dilution. The results of these tests are set forth in accompanying TableIX.

22. -A steroid according to Formula X of claim 21 wherein R is and R" ismethylene or ethylene.

23. A steroid according to claim 22 wherein R is methoxy and each of Zand Z is a single bond.

24. A steroid according to Formula X of claim 21 selected from those ofthe formula:

in which R R x, Z and are as defined therein.

25. A steroid according ,to claim 24, "wherein R is methoxy.

26. A quaternary ammonium steroid selected from those of the followingFormulas X-III, XIV,'XV and XVI:

19 and (XVI) wherein R is the group in which m is a positive integer oftwo to seven; n is a positive integer of two to four; and x is apharmaceutically acceptable anion; R is x0 orthe group Rd? in which R ishydrogen, hydroxy and pharmaceutically acceptable esters and ethersthereof, acetyl or lower alkyl of one to ten carbon atoms; 10 R ismethyl or ethyl; and

Z is a carbon-carbon single bond or a carbon-carbon double bond betweenC22 and C23.

References Cited UNITED STATES PATENTS 3,413,285 11/1968 Tsatsas260-239.5

ELBERT L. ROBERTS, Primary Examiner U.S. Cl. X.R.

